Opiate receptors 51 years on

It seems like only yesterday that Candace Pert found the morphine receptor  as a graduate student at Johns Hopkins 2 years after graduating BrynMawr at 24 and getting (academically) screwed by Solomon Snyder, who may have a department named after him at Hopkins but who will never get the Nobel prize.

Things seemed so simple back then, and the route to the nonaddicting opiate seemed clear.  Well its 51 years and counting since 1972 and things have become incredibly complicated as we learn more and more about our opiate receptors (there are 4).

A marvelous review is available (if you have a subscription) replete with multiple cryoEM structures of multiple receptors with multiple ligands at  resolution approaching amino acid size (better than 3.5 Angstroms).  Cell vol. 186 pp. 5203 – 5219 ’23.

Just in terms of combinatorial size, the possibilities are quite large.

There are 4 types of G Protein Coupled Receptors (GPCRs) binding opiate peptides — mu, delta, kappa and nociceptive.  Although we have all sorts of small molecule drugs binding to them (morphine, heroin, fentanyl and worse), their natural ligands in our brains are protein fragments (of which there are 20) derived from 4 precursor proteins) — so that’s 80 possibilities there.

To get anything done inside the brain, each of the 4 GPCRs binds to G proteins of the G(i/o) class of which there are 7.

But wait, there’s more.  All good things come to an end, and to stop signaling the intracellular part of the GPCR is phosphorylated by the awfully named GRKs (G receptor kinases) of which there are 7.

Once the G proteins are phosphorylated, they leave the intracellular part of the GPCR, and another protein (arrestin) binds to the same area of the GPCR.  There are 4 known arrestins.

The possibilities are multiplicative since they’re independent — so its

20 x 4 x 7 x 7 x 4 = 15,680 different possible interactions.

For a long time it was thought that arrestins terminated opiate peptide signaling, dragging the receptor inside the cell and schlepping it to either the lysosome or the proteasome where it was then destroyed by proteolysis.

Not so.  After arrestin binding, the opiate receptors can be found in endosomes where they can continue to signal, so we’re not really sure just what the effects of the arrestins are on opiate signaling.

Now let’s hear it for the biochemical ingenuity of plants.  Even the smallest opiate peptide (met-enkephalin) has 5 amino acids, far too large to insert itself in the 7 alpha helices of the GPCR crossing the cell membrane.  So they bind to the extracellular surface of the GPCR, particularly extracellular loop 2 (ICL2) which contains 21 amino acids.  Yet plants have figured out how to make small molecules like morphine which bind to the parts of the GPCR in the cell membrane.  It’s hard for me to see an evolutionary push (selection pressure) for them to do this.

Well that’s a rather broad overview of what’s in the paper, but there is much much more.  Reading it is like eating intellectual fruitcake — it is far too dense to be ingested and digested at one sitting.

Two further tidbits to whet your interest.  The paper contains a detailed discussion (with pictures of the structures) of why fentanyl is so much more potent than morphine.  But of course such things require as knowledge of organic chemistry.  The benzene ring of Fentanyl engages in direct pi pi electron interactions with the toggle switch amino acids tryptophans #295 and #328 (W295 and W328).  Benzene doesn’t contain an isolated benzene moiety.  Also the phenylethyl group of Fentanyl interacts hydrophobically with a minor binding pocket of morphine found between transmembrane 2 (TM2) and TM3.     Meat and drink for an old organic chemist such as yours truly.

Tidbit #2.  On activation of the opiate GPCR, there is inward movement of TM5 and outward movement of TM6 along with clockwise rotations of TM6.  This is initiated by ligand reconstruction of the polar network in the binding pocket, the collapse of the sodium pocket and rearrangement of the proline, isoleucine and phenylalanine triad (all 3 found on 3 different transmembrane segments (TMs).

Put the multiple structures shown in the paper are thousands of times more instructive than the previous two paragraphs, confirming yet again the old adage.